Compensation
(An informal perspective)
Prelude: You are free to copy and distribute this document at
will--but please do so in its entirety, complete with figures. For information
on obtaining a hardcopy for this purpose, click here.
To reference this material, please include the WWW address as well as the
latest modification date: March 11, 1996.
Address any questions, comments, or suggestions to Mario
Roederer
Introduction. Compensation is probably the least understood process
accompanying flow cytometric analyses. Perhaps this is because it is often
described with the linear algebra elements needed for its computation, and
many of us have forgotten what linear algebra is, much less how to do it.
However, proper compensation is absolutely crucial for some aspects of flow
cytometry, one of which is antigen density measurements. Unfortunately,
because compensation is often misunderstood, and because there is so much
incorrect mythology about it floating around, many laboratories do not set
compensation properly.
In this "perspective," I will try to address the issues surrounding
compensation, attempt to dispel many of the myths, and hope that by the
end, that I have not confused anyone but rather have convinced them what
proper compensation is.
Please start with the quiz, and work through the perspective in order suggested.
The table of contents below will allow you to go directly to any page.
Start with the Quiz!
If you are really adventurous, you can navigate through a more detailed
(and more rigorous) discussion of compensation, complete with examples of
the math, examples of the pitfalls of poor compensation, etc. I recommend
this only after you have finished with the more informal, relaxed discussion...
To start the detailed discussion, click here.
Table of contents:
Introduction
Quiz
Importance of compensation
Proper compensation
Errors introduced by compensation
Practical compensation
Autofluorescence
Recipe
Postscript
References
List of Figures:
Figure 1a (quiz, part
1)
Figure 1b (quiz, part 2)
Figure 2: Compensation &
cell enumeration)
Figure 3: Errors introduced
by compensation)
Figure 4: Practical compensation)
Figure 5: Compensation & sensitivity
settings)
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CD ROM Vol 2 was produced by staff at the Purdue University Cytometry Laboratories
and distributed free of charge as an educational service to the cytometry community.
If you have any comments please direct them to Dr. J. Paul Robinson, Professor & Director, PUCL,
Purdue University, West Lafayette, IN 47907. Phone:(317) 494-0757; FAX (317) 494-0517; Web http://www.cyto.purdue.edu
EMAIL robinson@flowcyt.cyto.purdue.edu